The ultimate expression of the genetic makeup of a cell resides in the proteins of that cell. Although protein synthesis is a complex process requiring numerous proteins and nucleic acids, to a large extent the specificity is determined by the first step of the process - the activation and transfer of an amino acid to its respective transfer RNA. Although considerable information about the interaction between aminoacyl- tRNA synthetases and tRNA has been obtained, relatively little is known about the control of protein synthesis that may be exercised at the level of aminoacyl-tRNA formation. Our long term goal is the elucidation of the pathways, enzymes and regulatory factors which determine the levels of functional tRNA and aminoacyl-tRNA synthetases in cells, and their effect on the various cellular processes in which aminoacyl-tRNA participates. The studies we propose should provide essential information in this central area of macromolecular metabolism. The specific aims of this project are: 1) to elucidate the complete maturation pathway for E. coli tRNA precursors including all the processing intermediates and relevant processing enzymes; 2) to characterize and study exoribonucleases for mechanism of action and additional in vivo functions; 3) to characterize exoribonuclease genes and examine the regulation of RNase expression; 4) to analyze the role of the -C-C-A end of tRNA in vivo and to identify functional domains of tRNA nucleotidyltransferase; 5) to analyze the structure and function of the mammalian aminoacyl-tRNA synthetase complex, particularly as it exists in vivo; 6) to elucidate the structural and functional organization of the mammalian translation system in vivo, particularly as it relates to channeling of aminoacyl-tRNA; and 7) to examine the role of aminoacyl-tRNA synthetases in tRNA maturation and transport. In view of the central role of proteins in all cell functions, and the importance of aminoacyl-tRNA formation for the specificity of this process, an understanding of the mechanism and regulation of this step is of great importance. Elucidation of these aspects of protein synthesis in normal cells is a prerequisite to any understanding of abnormal situations which may occur under pathological conditions.